Bacterial infections are the number one cause of human deaths, killing ~6 million people each year worldwide. Even in developed countries, such as the US, bacterial infections are once again recognized as a significant threat to public health because of widespread, acquired drug resistance. ?-Lactam antibiotics such as penicillins and cephalosporins are among the most often used antimicrobial agents. The most prevalent mechanism of bacterial resistance to ?-lactam antibiotics is the production of ?-lactamases that are able to hydrolyze and thereby inactivate the drugs. Although clavulanic acid and sulbactam are available for inactivation of serine ?-lactamases, no inhibitors are available that are broadly active against the many distinct metallo-?-lactamases (MBL). MBLs have now been recognized as an emerging clinical threat in that these enzymes, unlike serine ?-lactamases, are able to hydrolyze essentially all ?-lactams, including carbapenems (e.g., imipenem) which are last resort drugs for several multidrug resistant Gram-negative bacterial infections. In addition, many MBLs (e.g., IMPs and VIMs) are encoded by transferable metallo-?-lactamase genes on plasmids that have disseminated quickly worldwide. Some multidrug resistant strains of Pseudomonas and Acinetobacter spp. have already demonstrated significant resistance against imipenem due to MBL genes and there are few options available to treat these infections. However, due to low amino acid sequence homology among MBLs, the spectrum of activity of current inhibitors varies considerably among enzymes. It is apparent that a pressing need exists to design and develop novel inhibitors that have broad and potent activity against MBLs. The successful development of such inhibitors would offer new treatment options for epidemic drug resistant Gram-negative bacterial infections. For Specific Aim 1, we will design and develop a series of compounds based on a thiazolidine MBL inhibitor, which is our most potent compound in Preliminary studies and, importantly, meets both requirements of our two hypotheses: (1) the compound has a strong Zn(II) chelating group and (2) mimics the structure of penicillin. The activity of these compounds against a number of MBLs will be tested and (quantitative) structure activity relationships (SAR) be analyzed and used to design compounds with improved activity. The ability of the novel MBL inhibitors to restore the susceptibility of ?-lactam resistant bacteria will also be tested. In addition, x-ray crystallographic studies of metallo-?-lactamases, IMP-1 and Bla2, complexed with the novel inhibitors will be performed. For Specific Aim 2, another series of compounds will be designed and developed based on the second most potent compound identified in preliminary studies. In addition, based on the two hypotheses stated above, we propose to design and synthesize novel bicyclic compounds that not only closely mimic the structures of ?-lactams, but have a known Zn2+-binding group. PUBLIC HEALTH RELEVANCE: Many bacteria have now become resistant to carbapenems, a class of penicillin-like antibiotics used as last resort drugs, because they have acquired a protein called metallo-beta-lactamase. The proposed research is designed to lead to new potential adjuvant antibiotics that can inhibit the activity of this protein and thus restore the susceptibility of these bacteria to carbapenems.